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Clinic freeze-preservation protective solution composition for umbilical cord mesenchymal stem cells and application thereof

A technology of mesenchymal stem cells and protective solution, applied in the field of biomedicine, can solve problems such as difficulty in effectively maintaining the viability of mesenchymal stem cells, unknown components, and difficulty in large-scale application, and achieves good clinical application prospects, simple preparation methods, and stability sexual effect

Inactive Publication Date: 2017-07-28
安徽瑞杰赛尔生物科技有限公司
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

It can be seen from Table 1 of the examples of this patent document that after the cryopreservation solution I and II respectively freeze the cells for 1, 2, and 3 months and then recover and detect them, the cell viability of the freezing solution I is 96%, 92.6%, and 89%, its cell viability reduction rate is 3.5% / month, and the cell viability rate of cryopreservation solution II is 97.4%, 95% and 94.7% successively, and its cell viability reduction rate is 1.35% / month. Although the cryopreservation solution can be directly infused intravenously, the cell viability is maintained for a short time
Moreover, the mesenchymal stem cell cryopreservation solution contains human AB plasma, a large amount of which is unknown and has individual differences, so the standardization and stability of cell cryopreservation cannot be guaranteed, and there are potential safety hazards for allogeneic transfusion
In addition, there are few sources of healthy plasma, and the cost is high, making it difficult to apply on a large scale
[0007] In summary, the cell cryopreservation solution in the prior art is difficult to effectively maintain the viability of mesenchymal stem cells, and does not meet the conditions for clinical use

Method used

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  • Clinic freeze-preservation protective solution composition for umbilical cord mesenchymal stem cells and application thereof
  • Clinic freeze-preservation protective solution composition for umbilical cord mesenchymal stem cells and application thereof
  • Clinic freeze-preservation protective solution composition for umbilical cord mesenchymal stem cells and application thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0072] This embodiment relates to the preparation of a clinical cryopreservation protection solution for umbilical cord mesenchymal stem cells, which specifically includes the following steps:

[0073] 1) As shown in Table 1, the formulation of the frozen storage solution is mixed with compound dextran 40 injection, 20 wt% human serum albumin and DMSO, and the density is 5×10 6 / mL of umbilical cord mesenchymal stem cell suspension.

[0074] Table 1. Freezing solution formula table

[0075]

[0076] Control group: Mix 40 mL of LG-DMEM medium (40%) with 50 mL of fetal bovine serum (50%), add umbilical cord mesenchymal stem cells until the cell density is 5×10 6 / mL, then add 10mL DMSO. The formula of the control group was the commonly used mesenchymal stem cell cryopreservation solution in China.

[0077] 2) Detection of the viability of umbilical cord mesenchymal stem cells after resuscitation

[0078] Take the cells before cryopreservation to detect the cell viability;...

Embodiment 2

[0087] This embodiment relates to a method for preparing a cryopreservation solution for cryopreservation of umbilical cord mesenchymal stem cells, comprising the following steps:

[0088] 1) Take fresh newborn umbilical cord under sterile conditions, cut the umbilical cord into 1-2cm umbilical segment, remove the blood vessels, wash the blood stains with D-PBS, and then cut into 0.125cm 3 organization block. Use sterile tweezers to pick up the tissue pieces and spread them evenly in a 10cm petri dish. 37°C, 5% CO 2 Incubate for 30 min in the incubator to make the tissue pieces stick to the wall of the culture dish. Add 10 mL of complete culture medium dropwise to each Petri dish (pay attention to dripping carefully along the wall of the dish, so as not to cause impulse tissue blocks). 37°C, 5% CO 2 The culture was continued, and half of the medium was changed after 5 days (d6). After another 5 days, while removing the tissue block, half the amount of medium was changed a...

Embodiment 3

[0093] This example relates to the detection of cell characteristics after one year of cryopreservation of umbilical cord mesenchymal stem cells in clinical cryopreservation protection solution

[0094] Configure the cryopreservation solution according to the optimal formula 3 in Example 1 and cryopreserve another batch of umbilical cord mesenchymal stem cells. After one year of cryopreservation, after recovery in a 37°C water bath, the cell morphology, growth curve, adipogenic and osteogenic differentiation were performed. , and surface marker detection. Control group is the same as embodiment 1

[0095] 1) Observation of cell morphology

[0096] After recovery, add the cells to a 10cm dish and add complete medium at 37°C, 5% CO 2 After 24 h of culture, the morphology of cells in each group was observed under microscope.

[0097] 2) Cell growth curve

[0098] Cells before cryopreservation, cryopreservation solution of the present invention cryopreservation cell injection,...

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Abstract

The invention provides a clinic freeze-preservation protective solution composition for umbilical cord mesenchymal stem cells and application thereof. The protective solution composition comprises the following components: compound dextranum 40 injection, human serum albumin and DMSO. Compared with the prior art, the composition has the beneficial effects that the clinic freeze-preservation protective solution for umbilical cord mesenchymal stem cells can main the activity and biology activity of the mesenchymal stem cell for a long time, and does not contain unsafe components such as cell culture mediums, fetal calf serum and human plasma. All ingredients comprise clinical drug compound dextranum 40 injection, 20% human serum albumin or clinically used (dimethyl sulfoxide (UPS clinical level)), and the stability of the clinically standardized mesenchymal stem cell freeze-preservation system can be guaranteed while the umbilical cord mesenchymal stem cells for cryopreservation resuscitation meet the clinic using requirement. The application is safe, effective and simple in preparation method, and has good clinical application prospect.

Description

technical field [0001] The invention relates to a clinical cryopreservation protection solution composition for umbilical cord mesenchymal stem cells and an application thereof, belonging to the technical field of biomedicine. Background technique [0002] Mesenchymal stem cells (MSCs) are a type of pluripotent stem cells derived from mesoderm and ectoderm, which have the potential of self-renewal and multilineage differentiation. Studies have shown that mesenchymal stem cells can differentiate into adipocytes, smooth muscle cells, osteoblasts, chondrocytes, liver cells, nerve cells, glial cells, skin cells and other cell types, and have a good ability to repair damage. Mesenchymal stem cells were first discovered in the bone marrow and are mainly involved in the directional homing of bone marrow hematopoietic stem cells and the maintenance of homeostasis. In recent years, studies at home and abroad have found that mesenchymal stem cells not only have multi-directional diff...

Claims

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Application Information

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IPC IPC(8): A01N1/02
CPCA01N1/0221
Inventor 朱灏
Owner 安徽瑞杰赛尔生物科技有限公司
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